The present invention concerns the field of diagnostic measurement of nitric oxide in orally exhaled air and in particular how to avoid the influence of nitric oxide produced in the oral cavity in these measurements. Consequently, a composition, method and device for this purpose are disclosed.
It has been shown that increased NO levels in the orally exhaled breathing air is an indication of an inflammatory condition in the airways. A system and method for the determination of NO levels in orally exhaled air for diagnosis of inflammatory conditions is described, for example in WO 95/02181; Alving, K. et al. and WO 93/05709; Gustafsson, L. E.
Nitric oxide is normally produced enzymatically by constitutive NO synthases in, e.g. nerves and endothelial cells; these enzymes yield the relatively small amounts of NO involved in physiological regulation of nerve transmission and vascular tone. In contrast, the inducible NO synthase found, e.g. in activated white blood cells and airway epithelium produces NO at a high rate. It was recently shown that nitric oxide is also produced through a second route, i.e. through reduction of nitrite. Nitrite is present in body fluids such as saliva and urine, in amounts depending on the individual""s diet and health. For example, a diet rich in nitrate (such as certain vegetables) will result in high nitrate levels in saliva and urine. The bacteria normally present in the oral cavity will reduce salivary nitrate to nitrite.
The oral cavity or cavum oris is generally considered to comprise the cavity from the lips to epiglottis and this definition will be applied in the following description.
Further reduction of salivary nitrite to nitric oxide occurs normally in the acidic and reducing environment of the stomach (Lundberg et al. 1994, Gut, 35:1543-46). It is worth noting, that saliva contains relatively high amounts of ascorbic acid, the reducing capacity of which will enhance the production of NO from nitrite. Levels encountered in orally exhaled air can be in the interval of 30 to 50 pmol/s for healthy subjects and about 60 to 100 pmol/s for asthmatic patients. In healthy persons, the ingestion of nitrate rich food and the NO synthesis in the oral cavity may under certain conditions result in levels in the order of magnitude of 70 pmol/s. This is clearly in the asthmatic range, which makes this a potential source of error and misinterpretation of the results.
In Nature Medicine, vol. 1, no. 6, June 1995, Duncan et al. report their studies of nitric oxide generation in the oral cavity including experiments, where healthy volunteers have used an antibacterial mouthwash and an ascorbic acid mouthwash. Prior treatment with ascorbic acid caused an marked elevation in NO production and an initial reduction in salivary nitrite concentration. Conversely, prior treatment with antiseptic mouthwash (Eludril(copyright)) caused an initial reduction in NO production and a reduced salivary nitrite production. The results indicate, that NO synthesis was inhibited by 36xc2x110%. The authors are however focused on showing the beneficial effects of NO production and have not addressed the problems associated with this in relation to diagnostic measurements.
Surprisingly, the contribution of NO from the oral cavity may, under certain circumstances, be high enough to cause errors in diagnostic measurements, e.g. measurements according to WO 95/02181; Alving, K. et al. or WO 93/05709; Gustafsson, L. E. The consequences may be that a healthy person exhibits NO values corresponding to values of a person having asthma and an asthmatic, showing increased values may run the risk of being prescribed unnecessarily high doses of steroids or other anti-inflammatory medication. Obviously, with the development of this new concept of detection and diagnosis, which presently gains wider acceptance, there is an evident need for a reliable method and/or a safe composition which eliminates this possible source of error.
The present invention discloses a method and composition according to the attached claims, which for the first time substantially reduces or eliminates the possible source of error due to orally produced NO when measuring orally exhaled NO, and furthermore, does so in a practical, easy and reproducible way.